title: Chorein sensitivity of actin polymerization, cell shape and mechanical stiffness of vascular endothelial cells.
abstract: OBJECTIVE
  Endothelial cell stiffness plays a key role in endothelium-dependent control of vascular tone and arterial blood pressure. Actin polymerization and distribution of microfilaments is essential for mechanical cell stiffness. Chorein, a protein encoded by the VPS13A gene, defective in chorea-acanthocytosis (ChAc), is involved in neuronal cell survival as well as cortical actin polymerization of erythrocytes and blood platelets. Chorein is expressed in a wide variety of further cells, yet nothing is known about the impact of chorein on cells other than neurons, erythrocytes and platelets. The present study explored whether chorein is expressed in human umbilical vein endothelial cells (HUVECs) and addressed the putative role of chorein in the regulation of cytoskeletal architecture, stiffness and survival of those cells.
  
  
  METHODS
  In HUVECs with or without silencing of the VPS13A gene, VPS13A mRNA expression was determined utilizing quantitative RT-PCR, cytoskeletal organization visualized by confocal microscopy, G/F actin ratio and phosphorylation status of focal adhesion kinase quantified by western blotting, cell death determined by flow cytometry, mechanical properties studied by atomic force microscopy (AFM) and cell morphology analysed by scanning ion conductance microscopy (SICM).
  
  
  RESULTS
  VPS13A mRNA expression was detectable in HUVECs. Silencing of the VPS13A gene attenuated the filamentous actin network, decreased the ratio of soluble G-actin over filamentous F-actin, reduced cell stiffness and changed cell morphology as compared to HUVECs silenced with negative control siRNA. These effects were paralleled by a significant decrease in FAK phosphorylation following VPS13A silencing. Moreover, silencing of the VPS13A gene increased caspase 3 activity and induced necrosis in HUVECs.
  
  
  CONCLUSIONS
  Chorein is a novel regulator of cytoskeletal architecture, cell shape, mechanical stiffness and survival of vascular endothelial cells.
authors.affiliation: Department of Physiology, University of Tübingen, Tübingen, Germany.
authors.forename: Ioana
authors.initials: I
authors.lastname: Alesutan
authors.forename: Jan
authors.initials: J
authors.lastname: Seifert
authors.forename: Tatsiana
authors.initials: T
authors.lastname: Pakladok
authors.forename: Johannes
authors.initials: J
authors.lastname: Rheinlaender
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Lebedeva
authors.forename: Syeda T
authors.initials: ST
authors.lastname: Towhid
authors.forename: Christos
authors.initials: C
authors.lastname: Stournaras
authors.forename: Jakob
authors.initials: J
authors.lastname: Voelkl
authors.forename: Tilman E
authors.initials: TE
authors.lastname: Schäffer
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
mesh_terms: D000199:Actins / Q000378:metabolism*; D053148:Caspase 3 / Q000378:metabolism; D048430:Cell Shape; D003599:Cytoskeleton; D051416:Focal Adhesion Protein-Tyrosine Kinases / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D018625:Microscopy, Atomic Force; D008855:Microscopy, Electron, Scanning; D009336:Necrosis; D010766:Phosphorylation; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D000199:Actins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C429654:VPS13A protein, human; D033921:Vesicular Transport Proteins; D051416:Focal Adhesion Protein-Tyrosine Kinases; D053148:Caspase 3